Recombinant C1 beta-glucosidase for production of sugars from cellulosic biomass

• US 9,175,324 B2
• Filed: 11/03/2014
• Issued: 11/03/2015
• Est. Priority Date: 09/30/2009
• Status: Active Grant

• Current Assignee: Codexis, Inc.
• Original Assignee: Codexis, Inc.
• Inventors: Clark, Louis A., Baidyaroy, Dipnath, Szabo, Lorand
• Primary Examiner(s): RAMIREZ, DELIA M
• Application Number:
  US14/531,727
• Publication Number:
  US 20150050702A1
• Time in Patent Office: 365 Days
• US Class Current: 1/1
• CPC Class Codes: C12N 9/2445 Beta-glucosidase (3.2.1.21)
• CPC Class Codes: C12P 19/02 Monosaccharides 2-ketogulonic acid C12P7/60
• CPC Class Codes: C12P 19/14 produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase , e.g. by cellulase, hemicellulase
• CPC Class Codes: C12Y 302/01021 Beta-glucosidase (3.2.1.21)

Abstract

The invention relates to expression of a recombinant C1 β-glucosidase. The invention also provides methods for producing a fermentable sugar from cellobiose by contacting cellobiose with a recombinant host cell comprising a polynucleotide sequence encoding C1 β-glucosidase, operably linked to heterologous promoter, under conditions in which β-glucosidase is expressed and secreted by the cell and the cellobiose is enzymatically converted by said β-glucosidase to glucose. Methods of the invention may be used for conversion of a biomass substrate to a fermentable sugar, and ultimately to ethanol or other biofuel.

Claims

1. A method for producing glucose from cellobiose, comprising contacting cellobiose with a recombinant cell comprising a polynucleotide encoding a recombinant β

-glucosidase, wherein said polynucleotide comprises SEQ ID NO;

9 and is operably linked to a heterologous promoter, under conditions in which said β

-glucosidase is produced by the cell and said cellobiose is enzymatically converted by said β

-glucosidase to glucose.

2. The method of claim 1, wherein the recombinant cell is a fungal cell.

3. The method of claim 2, wherein the recombinant cell is a Myceliophthora thermophila cell.

4. The method of claim 1, wherein the heterologous promoter is the M. thermophila C1 cellobiohydrolase 1a (CBH1a) promoter.

5. The method of claim 1, wherein the recombinant cell further comprises at least one additional cellulase enzyme.

6. The method of claim 5, wherein said additional cellulase enzyme is selected from a cellobiohydrolase, an endoglucanase, and a β

-glucosidase.

7. The method of claim 5, wherein said additional cellulase enzyme is a heterologous enzyme.

8. A method of converting a biomass substrate to a fermentable sugar, comprising combining a recombinant host cell with a biomass substrate, wherein the recombinant host cell comprises a polynucleotide encoding a recombinant β

-glucosidase, wherein said polynucleotide comprises SEQ ID NO;

9 and is optionally operably linked to a heterologous promoter, under conditions suitable for the production of the fermentable sugar.

9. The method of claim 8, wherein said biomass substrate comprises cellobiose and the fermentable sugar is glucose.